Clostridium difficile is a Gram-positive bacterium that is known to be the cause of enteric diseases. It is the leading cause of antibiotic-associated diarrhea and pseudomembranous colitis (Knoop, F. C.; Owens, M.; Crocker, I. C.; Clin. Microbiol. Rev. 6, 1993, 251-265). The frequency and severity of outbreaks associated with C. difficile has increased in recent years (Pepin, J.; Valiquette, L.; Alary, M. E.; Villemure, P.; Pelletier, A.; Forget, K.; Pepin, K.; Chouinard, D.; JAMC. 171, 2004, 466-472). Most ingested vegetative C. difficile cells are destroyed by the acidic environment present in the stomach. Spores, however, can survive this feat and upon exposure to bile acids can germinate in the small bowl. The decreased levels of normal microbial cells in the intestines due to medical treatments such as antibiotics use and chemotherapy, allow C. difficile to proliferate.
In humans, C. difficile-associated diarrhea (CDAD) is the most commonly diagnosed cause of hospital-associated and antimicrobial-associated diarrhea. Risk of CDAD has traditionally been higher among elderly patients and those that have undergone hospitalization, gastrointestinal surgery or were exposed to antibiotics. In the United States, the estimated number of cases of C. difficile-associated disease exceeds 250,000 per year (Wikkins T D and Lyerly D M, 2003, J. Clin Hasp Infect 48:81), with total additional health care costs approaching US $1 billion annually (Kyne L, et al., 2002, Clin Infect Dis 34:346-353).
In the past five years an unexpected increase in the incidence of CDAD has been observed. This has also been associated with higher rates of severe CDAD, treatment failure and death. Severe cases are being more frequently identified in younger patients and those without traditional risk factors. Much of this change has been associated with international dissemination of an outbreak clone, designated ribotype 027 (also known as North American pulsotype 1 (NAP1) and BI). Prevention of C. difficile is based on patient isolation, improved sanitation, improved infection control and antimicrobial restriction, all of which are associated with high healthcare costs. In addition, prophylactic use of antibiotics has been used for the prevention of infection; however, it led to an increase in the incidence of disease. Treatment of C. difficile infections is also problematic since the response to metronidazole, the main first-line treatment, is becoming unpredictable. Vancomycin, the alternative choice, is expensive and its use raises concern about emergence of vancomycin-resistant enterococci and other vancomycin-resistant organisms.
CDAD is also an important problem in many animal species such as horses and pigs. It may also be a cause of disease in other species. There is concern that C. difficile may be transmissible from animals to humans because the types of C. difficile isolated from animals are often the same as those found in people, including the outbreak strain ribotype 027. This concern has increased based on the finding of C. difficile in retail meat samples.
The reported increasing incidence of CDAD, its recurrence rates, and its impact on morbidity and mortality, as well as the costs associated with treatment and appropriate isolation procedures to limit its spread make clear the need for effective prevention approaches of CDAD.
One particular strain, designate ribotype 027 or NAP1 has emerged as an important cause of sporadic and epidemic disease internationally. Serious outbreaks with high morbidity, high mortality, poor response to treatment and high relapse rates have been reported. This strain produces 3 main toxins: toxin A, toxin B and CDT (binary toxin). It also has a deletion in a purported toxin regulating gene that appears to increase toxin production, at least in vitro (Just, I.; Selzer, J.; Wilm, M.; von Eichel-Streiber, C.; Mann, M.; Aktories, K.; Nature. 375, 1995, 500-5033). This spore-forming bacterium is found to resist phagocytosis by cell surface polysaccharides.
Poxton and Cartmill (Poxton, I. R; Cartmill; T. D. J Gen Microbiol. 1982, 128, 1365-1370) described the sugar composition of two preparations extracted from C. difficile strain NCTC 11223. The material obtained by NaOH treatment of cells was observed to contain glucose, mannose, galactosamine and phosphate, and the other, extracted by phenol treatment of cells, contained glucose, glucosamine, phosphate and fatty acids. In the same work, it was also observed that both preparations cross-reacted with Clostridium sordellii antiserum.
There is a growing need to develop a vaccine for humans and animals against C. difficile infection to prevent CDAD or prevent recurrence. Additionally, vaccination is needed in animals to prevent animal disease and to reduce shedding of C. difficile so as to reduce the risk of zoonotic transmission.